1. Field of the Invention
The present invention concerns immune responses initiated by the recognition of a peptide:MHC complex on the surface of antigen presenting cells by T-cells. The present invention also concerns immune responses initiated by the binding of a Signal-2 moiety to its complement protein on the surface of an antigen presenting cell. More particularly, the present invention concerns the immune responses initiated by the recognition of the peptide:MHC by the T-cell and by the binding of a Signal-2 moiety to its complement protein. Still more particularly, the present invention concerns the modification of the typical immune response generated by a particular individual in response to this binding. Most particularly, the present invention concerns the conjugation of peptides derived from the peptide portion of the peptide:MHC complex to the preferred Signal-2 moiety in order to modify or shift a given immune response from type-1 to type-2 or from type-2 to type-1. This may include specific phenotypes of regulatory T-cells including suppressor T-cells.
2. Description of the Prior Art
Autoimmune diseases are characterized by the activation of T-cells against self-antigens. These T-cells then destroy cells presenting these antigens. For example, insulin-dependent diabetes mellitus (IDDM, also called Type-I diabetes) is characterized by the activation of T-cells against the insulin-producing cells of the pancreas and their subsequent destruction by these T-cells. The diseases and conditions associated with autoimmune responses are strongly associated with specific subtypes (alleles) of cell surface proteins called major histocompatability complex (MHC) class II molecules. MHC molecules bind fragments (peptides) of proteins from infectious agents, allergens, and selfproteins, and this MHC:peptide complex is the structure that T-cells recognize with their receptor (called the T-cell receptor, or TCR). The MHC:peptide complex is displayed on the surfaces of other cells of the immune system (i.e., B cells, dendritic cells and macrophages) which are called antigen presenting cells (APC). In order for an immune response to ensue, the major regulatory cell of the immune system, the undifferentiated T-cell, must be presented with small breakdown products (peptides) of the foreign invader. This presentation occurs on the surface of the APC. The T-cell must then interact with the APC, and this interaction stimulates the T-cell to divide and differentiate to produce molecules that attack, either directly or indirectly, cells displaying the same or highly similar MHC:peptide complex. It is well known that the genes that encode the MHC molecules are extremely variable within the species, and the different MHC alleles prefer to bind some peptides over others. Along with other genetic and environmental factors, the existence of different MHC alleles helps to explain why some members of a species develop conditions such as autoimmune diseases, allergies, asthma, and even certain infectious diseases, while others remain seemingly unaffected, or immune, to the same substances. Other differences arise because cell surface proteins distinct from the peptide:MHC complex must also bind to specific receptors on the T-cell. These other protein:protein pairs at the interface of the T-cell and APC membranes provide a costimulatory signal, known as Signal-2 which, along with the signal generated by the TCR recognition of the MHC:peptide complex (known as Signal-1), initiates an immune response.
A defining stage of the immune response is the differentiation of CD4+ T-cells into either type-1 helper T-cells (TH1 cells) or type-2 helper T-cells (TH2 cells) as a result of the two signals. These two subtypes of TH cells and the regulatory network of cells that they selectively activate are well-known correlates of human health conditions and disease states. Differentiation into TH1 cells results in predominantly cell-mediated immunity while differentiation into TH2 cells results in predominantly humoral immunity. Each of these immunity types helps to protect the body against different types of invasion. Type-1 immunity protects the body against intracellular pathogens such as bacteria, but is also implicated in organ-specific autoimmune diseases. Type-2 immunity is important for protection against extracellular parasites, but is associated with allergic reactions as well. Development of TH1 cells is driven by a cytokine called interleukin-12, which is produced by immune cells known as macrophages and dendritic cells. Interleukin-12 induces or stimulates the naive T-cell (CD4+ T-cells) to produce interferon-γ (IFN-γ) and interleukin-2 (IL-2). These two cytokines (IL-2 and IFN-γ) are involved in classic cell-mediated functions such as clonal expansion of cytotoxic T-lymphocytes (CTLs), macrophage activation, and class switching to IgG isotypes that mediate complement lysis of sensitized cells. Commitment to a TH1 immune response is enhanced by the presence of IFN-γ which up-regulates expression of the interleukin-12 (IL-12) receptor while inhibiting the development of TH2 cells. TH2 immunity results from the production of interleukin-4 (IL-4) by the naive T-cell. IL-4 induces TH2 development and the subsequent production of interleukins-4 (IL-4), -5 (IL-5), -10 (IL-10), and -13 (IL-13). IL-4 also operates to down-regulate expression of the IL-12 receptor on developing cells, thereby inhibiting TH1 development and helping undifferentiated T-cells to commit to TH2 cell development. Additionally, IL-4 and IL-5 are known to activate B cells and switch to neutralizing antibody (IgG1 in the mouse) and IgE, the initiator of immediate hypersensitivity.
In order for either of these immune pathways to be activated, a two-signal mechanism is required to fully activate the T-cell. Signal-1 (S-1) occurs when the T-cell antigen receptor (TCR) recognizes the peptide:MHC-II complex on the surface of an antigen presenting cell (APC). This first signal passes through the T-cell receptor and initiates a cascade of tyrosine phosphorylation/dephosphorylation events mediated by kinases and phosphatases and leads to the activation of Ca++ flux, nuclear factor of activated T cells (NF-AT) and NFκB transcription factors. These factors enter the nucleus of the T-cell and bind to promoters of genes responsible for effector functions. Signal-2 (S-2) arises from the binding of Signal-2 receptors to their ligands on the surface of an APC. Signal-2 receptors include CD28 and its ligand B7 as well as LFA-1 and its ligand ICAM-1. When a Signal-2 receptor and its ligand form a complex at the interface between the T-cell and APC receptor membranes, a series of signaling events occur. These events include serine/threonine phosphorylation/dephosphory-lation and activation of guanine nucleotide exchange factors that activate adapter proteins with GTPase activity. These signaling events activate a separate set of transcription factors. The signal delivered through the CD28:B7 complex is distinct from that delivered from the ICAM-1:LFA-1 complex, particularly with respect to the differentiation of CD4+ T-cells into TH1 versus TH2 effector populations. When the predominant binding occurs between LFA-1 and ICAM-1, the CD4+ T-cell differentiation favors TH1 cells which are abundant producers of IL-2 and IFNγ, the preeminent initiators of inflammatory immune responses including delayed-type hypersensitivity (DTH), immunity to intracellular pathogens, and several autoimmune diseases. When the predominant binding occurs between CD28 and B7, the CD4+ T-cells differentiate into TH2 cells. In contrast to TH1 cells, TH2 cells do not produce abundant IL-2 or IFNγ cytokines, but instead release the mediators of immediate-type hypersensitivity such as allergy and asthma, i.e., IL-4, IL-5, IL-10, and IL-13. Thus, the ability to manipulate the relative contribution of the complex providing the second signal has a profound effect on the type of immune response that is elicited against a given self-tissue antigen.
The associations between the TCR and APC occur at a specialized junction or interface between the TCR and the APC called the immunological synapse. An immune synapse is depicted schematically in FIG. 1. This immune synapse can be defined as the organized structure of activation molecules that assemble at the interface between the T-cell and the APC. Like a synapse in the nervous system, the immune synapse is a close association between cellular membranes. In order for an immune response to ensue, the major regulatory cell of the immune system, the undifferentiated T-cell must be presented with small breakdown products (peptides) of the foreign invader. In an unactivated T-cell, TCR and adhesion molecules are dispersed randomly on the T-cell membrane. The formation of the immunological synapse is an active and dynamic mechanism that allows T-cells to distinguish potential antigenic ligands. The immunological synapse consists of a central cluster of T-cell receptors surrounded by a ring of adhesion molecules. The stable formation of the immune synapse requires adhesion molecules such as LFA-1 and the peptide-recognition receptor (TCR) to form a doughnut-like structure with the TCR on the inside and LFA-1 on the outside. During activation, the TCR and LFA-1 molecules pass by each other within the T-cell lipid bilayer during the formation of the doughnut-like structure (this process is called translocation). If these molecules do not translocate within the immune synapse then the T-cell signal is not fully received and a different program of gene activity may ensue within the T-cell. This can drastically effect the immune response, especially if the T helper cell deviates from a gene program that would lead to IFNγ release (TH1 cells and type-1 immunity) to a program that ultimately activates IL-4 production (i.e., TH2 cells and type-2 immunity).
In more detail, to activate the pathway leading to TH1 dominance, the TCR recognizes the peptide:MHC-II complex and sends Signal-1 to the T-cell. Additionally, LFA-1 binds to ICAM-1, and these molecules, along with the peptide:MHC-II complex, translocate to form the end-stage immune synapse. This leads to the effective expression of the CD40 ligand (CD154) by the uncommited TH cell. CD40 interaction (expressed on the antigen presenting cell) with its ligand generates NFκB up-regulation of the inflammatory cytokine, IL-12. IL-12 then binds to its receptor on the undifferentiated TH cell and initiates the TH1 program, including the up-regulation of the transcription regulators, Stat4 and Tbet. This leads to TH1 dominance against the autoantigen (e.g., glutamic acid decarboxylase, GAD65), which was initiated by the GAD65 peptide component of the TCR:peptide:MHC-II complex. For the pathway leading to TH2 dominance, the TCR can recognize the same peptide:MHC-II complex, thereby sending Signal-1. However, in this case, a weaker strength of Signal-1 and/or altered or blocked binding between Signal-2 moieties leads to an altered form of the end-stage immune synapse. Likely, this lower strength of Signal-1 or distinct participation of the LFA-1 second signal leads to this different result, i.e., dominant TH2 differentiation. For example, the altered immune synapse can dictate that the CD40 ligand is not expressed and IL-12 is therefore not released by the APC. This pathway is schematically represented in FIG. 2. Here, IL-4 appears to accumulate, thereby leading to the up-regulation of Stat6 and GATA-3 within the T-cell and hence commitment to a TH2 pattern of differentiation.
A major goal of modern applied immunology is to be able to switch from TH1-dominant immunity (e.g., as seen in autoimmune diseases and transplant rejection) to TH2 responses against these same tissue antigens. In other cases, it would be extremely valuable to replace weak TH2 immunity with TH1 dominance leading to strong T-cell proliferation and the effective generation of cytotoxic T-cells (CTL). These cases may include chronic viral illnesses, like hepatitis-C and AIDS; and could include certain cancers like melanoma. Accordingly, what is needed in the art is modifiers of these immune responses so that type-2 immunity can be replaced with type-1 immunity or type-1 immunity can be replaced with type-2 immunity, as desired in order to combat different human disease states or health conditions.